1. Field of the Invention
Foot-and-mouth disease virus (FMDV) is responsible for one of the most devastating and contagious diseases in cattle and other cloven-hooved animals, affecting over 100,000 animals a year and resulting in significant economic loss. The disease occurs in many areas of the world outside the United States where vaccination programs have been largely effective. There are risks associated with the vaccines currently in use, however, and at present FMDV vaccines cannot be produced in the United States despite the continued threat of the introduction of this agent into the country. It is feared that the virus used to make vaccines could escape from containment and cause disease. Moreover, the failure to completely inactivate the virus during vaccine preparation has led to accidental outbreaks of infection. In addition, there is considerable antigenic variability among the various serotypes, thus some viruses may not be recognized by the vaccinated animals. Furthermore, frequent revaccination has been required in order to maintain protective immunity utilizing conventional vaccines containing virus attenuated by chemical inactivation (Bachrach, H. L. 1968. Annu. Rev. Microbiol. vol. 22, pp. 201-244). There is thus a strong incentive to develop an effective vaccine which eliminates the threat of infection due to the accidental outbreaks associated with vaccine production and administration. A new and safer genetically-engineered vaccine against FMDV which provides effective protection but is not infectious, and thus does not present the risk of causing accidental infections, is the subject of related patent application Ser. No. 08/418,716 now U.S. Pat. No. 5,612,040. This invention relates to a genetically-engineered cell line for the propagation of the mutant virus.
2. Description of the Related Art
In an effort to overcome the deficiencies of conventional virus vaccines, synthetic vaccines have been investigated. Identification of a flexible loop exposed on the virus surface as the main antigenic site of FMDV (site A) prompted the investigation of the use of various peptide fragments within site A to stimulate immunological responses. For example, the conserved tripeptide Arg-Gly-Asp (RGD) was evaluated for its ability to stimulate the production of neutralizing antibodies in rabbits or guinea pigs (Novella et al. 1993. FEBS Letters. vol. 330, no. 3, pp. 253-259).
Attempts to produce attenuated virus vaccines by genetic engineering were also carried out. Rieder et al. (1993. J. Virol. vol. 67, no. 9, pp. 5139-5145, herein incorporated by reference), for example, evaluated the role of the poly(C) tract found at the 5' end of the FMDV genome. Cardioviruses having shorter-than-natural poly(C) tracts had been shown to be dramatically attenuated; however, the poly(C) tract length of FMDV showed no effect on virulence when tested in mice.
A non-infectious form of the FMD virus consisting of a genetically-engineered receptor-binding site-deleted virus particle has been described in related U.S. Pat. No. 5,612,040, supra. The receptor binding site-deleted particle, which lacks the three amino acids of the tripeptide RGD in the capsid protein VP1 is not infectious for cells in culture or for animals; however, the RGD-deleted virion is able to induce a protective immune response in cattle U.S. Pat. No. 5,612,040, supra; McKenna et al. 1995. J. Virol. vol. 69, p. 5787). Since the virus cannot infect cells in culture or animals, production of the virus has been limited to the transfection of cells capable of allowing assembly of the mutant virus from the vector containing the mutant RNA, allowing RNA to replicate in the transfected cells and harvesting the mutant virus particles from the cell cultures. Although effective, it is expensive to produce commercial quantities of virus in this manner. Thus a means for the production of virus in larger quantities was needed.